Coral Composite Extract, Composition Including The Same And Method Of Producing The Same

ABSTRACT

The present invention relates to a coral composite extract, a composition including the same and a method of producing the same. The coral composite extract includes at least two briarane-type diterpenoid compounds from corals of Briareum violaceum, B. excavatum and B. stechei, thereby being applied as an effective ingredient of a skin external use composition, a cosmetic composition and a medicinal composition.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a Divisional Application of the U.S. applicationSer. No. 16/577,422, filed Sep. 20, 2019, which claims priority toTaiwan Application Serial Number 107145343, filed Dec. 14, 2018, whichis herein incorporated by reference in its entirety.

BACKGROUND Field of Invention

The present invention relates to a marine natural extract. Morespecifically, the present invention relates to a coral compositeextract, a composition including the same and a method of producing thesame.

Description of Related Art

The oceans cover about 70% of the Earth's surface and are the originswhere life is born and bred. In the evolution process, marine organismsproduce secondary metabolites that are different from terrigenousnatural compounds and have unique structures to be adapted to thestringent environments such as hypersaline, high-pressure and anoxicenvironments, and so on, so as to obtain advantages for survival andcompetition.

However, marine resources and bioecological environments suffer variouslevels of damages due to overexploitation and anthropogenic destructionof partial marine environment recently. How to make good use of marineresources is one of the research priorities under the premise ofecological conservation of marine environments.

Taiwan is an ideal place for further research focusing on the secondarymetabolites produced by corals since the coral systems in the sea areaaround Taiwan are well-developed and contain various species. Under thepoint of view, extensive Taiwanese experience to breed corals should beapplied not only in conserving the marine ecology and reducing the shockto the marine living resources, but also in developing naturally activecompounds to promote the application of marine living resources.

SUMMARY

Accordingly, one aspect of the present invention is to provide a coralcomposite extract obtained from a coral starting material by a crudeextraction step and a column chromatography step, and the coralcomposite extract includes at least two briarane-type diterpenoidcompounds.

In another aspect, the invention provides a coral composite extractobtained from a coral starting material originated from Briareumviolaceum by a crude extraction step and a column chromatography step,in which the coral composite extract includes excavatolide B (EXC-B) andEXC-Z.

In the other aspect, the invention provides a coral composite extractobtained from a coral starting material originated from B. excavatum bya crude extraction step and a column chromatography step, in which thecoral composite extract includes EXC-B and EXC-C.

In the other aspect, the invention provides a coral composite extractobtained from a coral starting material originated from B. stechei by acrude extraction step and a column chromatography step, in which thecoral composite extract includes brianolide (BR), briarenolide X (BR-X)and BR-W.

In the other aspect, the invention provides a composition including theaforementioned coral composite extract as an effective ingredient.

In the other aspect, the invention provides a method of producing thecoral composite extract, which includes performing a crude extractionstep and a column chromatography step on the coral starting material, soas to obtain the aforementioned coral composite extract.

According to the aforementioned aspect, the invention provides a coralcomposite extract. In an embodiment, the coral composite extract isobtained from a coral starting material by a crude extraction step and acolumn chromatography step, in which the coral starting material caninclude but be not limited to B. violaceum, B. excavatum and B. stechei.The aforementioned coral composite extract includes at least twobriarane-type diterpenoid compounds, and the briarane-type diterpenoidcompounds can be excavatolide B (EXC-B), EXC-C, EXC-Z, brianolide (BR),briarenolide X (BR-X), and BR-W, for example.

According to the aforementioned aspect, the invention further provides acoral composite extract. In one embodiment, the coral composite extractcan be obtained from a coral starting material originated from B.violaceum by a crude extraction step and a column chromatography step,in which the aforementioned coral composite extract can include EXC-Band EXC-Z, for example.

According to the aforementioned aspect, the invention further provides acoral composite extract. In one embodiment, the coral composite extractcan be obtained from a coral starting material originated from B.excavatum by a crude extraction step and a column chromatography step,in which the aforementioned coral composite extract can include EXC-Band EXC-C, for example.

According to the aforementioned aspect, the invention further provides acoral composite extract. In one embodiment, the coral composite extractcan be obtained from a coral starting material originated from B.stechei by a crude extraction step and a column chromatography step, inwhich the aforementioned coral composite extract can include at leasttwo of brianolide (BR), briarenolide X (BR-X), and BR-W.

According to the aforementioned aspect, the invention further provides acomposition applying the aforementioned coral composite extract as aneffective ingredient.

According to the aforementioned aspect, the invention further provides amethod of producing a coral composite extract. In one embodiment, themethod performs a crude extraction step with a first solvent on thecoral starting material to obtain a crude extract at first. In theaforementioned embodiment, the coral starting material can be B.violaceum, B. excavatum or B. stechei, for example, and the firstsolvent can include water, alcohol solvent, ester solvent, alkanesolvent, and ketone solvent.

Furthermore, a column chromatography step is performed on theaforementioned crude extract by conducting a gradient elution treatmentwith a chromatography column to obtain the coral composite extract. Inthe aforementioned embodiment, the gradient elution treatment can beconducted with a second solvent, in which the second solvent can includethe aforementioned ester solvent and/or alkane solvent, the coralcomposite extract can include at least two briarane-type diterpenoidcompounds, and the at least two briarane-type diterpenoid compound caninclude EXC-B, EXC-C, EXC-Z, BR, BR-X and BR-W.

In one embodiment of the invention, the aforementioned coral startingmaterial can be lyophilized.

In one embodiment of the invention, the aforementioned crude extractionstep can further include a combination of an ultrasonic treatment and/ora microwave extraction treatment.

In one embodiment of the invention, the aforementioned alcohol solventcan be methanol, ethanol, propanol and isopropanol, for example.

In one embodiment of the invention, the aforementioned ester solvent canbe ethyl acetate, isopropyl acetate, butyl acetate, methoxyethanolacetate and ethoxyethanol acetate, for example.

In one embodiment of the invention, the aforementioned ketone solventcan be acetone, for example.

In one embodiment of the invention, the ester solvent of theaforementioned second solvent can be ethyl acetate, for example, and thealkane solvent of the second solvent can be n-hexane, for example.

In one embodiment of the invention, a concentration gradient (volumeratio) of n-hexane to ethyl acetate of the second solvent can be 100:0to 0:100, for example, when conducting the aforementioned gradientelution treatment.

By applying the coral composite extract, the composition including thesame and the method of producing the same of the present invention, thecoral composite extract includes at least two briarane-type diterpenoidcompounds and can be added to the composition as an effectiveingredient.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention can be more fully understood by reading the followeddetailed description of the embodiment, with reference made to theaccompanying drawings as follows:

FIG. 1 is a partial flow chart showing the method of producing coralcomposite extract according to one embodiment of the present invention.

FIG. 2 is a partial flow chart showing the method of producing coralcomposite extract according to another embodiment of the presentinvention.

FIG. 3 is a partial flow chart showing the method of producing coralcomposite extract according to the other embodiment of the presentinvention.

FIG. 4 is a partial flow chart showing the method of producing coralcomposite extract according to the other embodiment of the presentinvention.

FIG. 5 is a bar chart of the extraction rate of the crude extractobtained by the crude extraction step in different ways according toTABLE 1.

DETAILED DESCRIPTION

Singular forms, “a”, “an”, and “the”, as mentioned in the presentinvention also cover plural references, unless otherwise stated in thecontext. A value range (such as 10% to 11% of A) includes its upper andlower limits (i.e. 10%≤A≤11%) if not otherwise stated specially; thevalue range, for which if no lower limit is defined (such as B lowerthan 0.2%, or B below 0.2%), can have a lower limit of zero (i.e.0%≤B≤0.2%). The aforementioned terms were used for illustrating andhelping understand the present invention rather than limiting the scopethereof.

The invention provides a coral composite extract obtained from a coralstarting material by a crude extraction step and a column chromatographystep, and the coral composite extract includes at least twobriarane-type diterpenoid compounds.

The “coral composite extract” mentioned in the present invention isobtained from the coral starting material by the crude extraction stepand the column chromatography step. In one embodiment, theaforementioned coral starting material can include but be not limited toBriareum violaceum, B. excavatum and B. stechei.

In the embodiment, the obtained coral composite extract can include atleast two briarane-type diterpenoid compounds. Here, the mentioned atleast two briarane-type diterpenoid compounds are the two briarane-typediterpenoid compounds with the highest amounts of the coral compositeextract instead of limiting the coral composite extract to only twobriarane-type diterpenoid compounds. In the aforementioned embodiment,the briarane-type diterpenoid compounds can be excavatolide B (EXC-B),EXC-C, brianolide (BR), briarenolide X (BR-X) and BR-W, for example.

In one example, the aforementioned coral composite extract can beobtained from the coral starting materials originated from B. violaceumby the crude extraction step and the column chromatography step, and thecoral composite extract includes EXC-B and EXC-Z.

In one example, the aforementioned coral composite extract can beobtained from the coral starting materials originated from B. excavatumby the crude extraction step and the column chromatography step, and thecoral composite extract includes EXC-B and EXC-C.

In one example, the aforementioned coral composite extract can beobtained from the coral starting materials originated from B. stechei bythe crude extraction step and the column chromatography step, and thecoral composite extract includes BR, BR-X and BR-W.

FIG. 1 is a partial flow chart showing the method 100 of producing coralcomposite extract according to one embodiment of the present invention.First, the coral starting material is provided as STEP 101 depicts. Inone embodiment, the aforementioned coral starting material can includebut be not limited to B. violaceum, B. excavatum and B. stechei. Theaforementioned coral starting material can optionally be pretreated withknown drying methods, such as atmospheric drying, vacuum drying,lyophilization, vacuum lyophilization, and so on, so as to removemoisture from the coral starting material without affecting anddestroying the amount as well as the activity of the coral compositeextract.

Next, as STEP 103 depicts, the crude extraction step is performed on thecoral staring material to obtain a crude extract, in which a firstsolvent is applied. In one embodiment, there is no limitation of thekind of the first solvent. However, the one having a polarity index P′value closer to a polarity index P′ value of the coral composite extractis the better. For example, a partition coefficient (log P) of EXC-B isabout 0.9 and an octand-water partition coefficient (Clog P) of EXC-B isabout 3.5568. Therefore, the polarity index P′ value of the firstsolvent can be 0.1 to 6.0, but 1.0 to 5.4 is better.

In the aforementioned embodiment, specific examples of the first solventcan include but be not limited to water, alcohol solvent, ester solvent,alkane solvent and ketone solvent. The alcohol solvent is short-chainalcohol, for example, alcohol having 1 to 3 carbon atoms, in which thespecific examples can be methanol, ethanol, propanol and isopropanol butnot allyl alcohol and propargyl alcohol. The ester solvent can includebut be not limited to ethyl acetate, methoxyethanol acetate andethoxyethanol acetate. The alkane solvent can include but be not limitedto n-hexane. The ketone solvent can include but be not limited toacetone, methyl ethyl ketone (MEK).

In the aforementioned embodiment, a solid-liquid ratio of theaforementioned coral starting material to the aforementioned firstsolvent is not limited. In one example, the solid-liquid ratio (g:mL) ofthe aforementioned coral starting material to the first solvent can be1:1 to 1:20, for example. However, 1:1 to 1:15 is better, and 1:1 to1:10 is much better.

In the aforementioned embodiment, the crude extraction step can beeither conducted by the solvent extraction treatment with the firstsolvent along or the solvent extraction treatment in combination with anultrasonic treatment and/or a microwave extraction treatment, or asupercritical extraction treatment with a mixture of the aforementionedalcohol solvent and a supercritical fluid. The aforementionedsupercritical fluid can be carbon dioxide, for example.

In general, a boiling point of the aforementioned first solvent is lowerthan a melting point of the coral composite extract (about 224° C.).Thus, it is better that the ultrasonic treatment and/or the microwaveextraction treatment are conducted under the temperature lower than theboiling point of the first solvent to perform the crude extraction step.In one specific example, the crude extraction step can be performedunder the temperature lower than 156° C., 145° C., 100° C., 78° C. or77° C. However, the temperature is not limited to the aforementionedexamples but depends on the extraction solvent.

In the aforementioned embodiment, there is no specific limit on how manytimes the crude extraction step is performed. Rather, the crudeextraction step can be performed once or repeated for many times toincrease the yield of the crude extract. In one example, the crudeextraction step can be repeated for once to 10 times, and preferablyonce to 6 times.

In the aforementioned embodiment, the coral starting material caninclude but be not limited to B. violaceum, B. excavatum and B. stechei.In one example, the coral starting material can be unprocessed orlyophilized.

However, as STEP 107 in FIG. 1 depicts, in one embodiment, after theaforementioned crude extraction step, the column chromatography step canbe performed by conducting the gradient elution treatment on the crudeextract with a chromatography column to obtain factions of the coralcomposite extract, as STEP 109 depicts.

In the aforementioned embodiment, the aforementioned chromatographycolumn can be a commercially available silica chromatography column, andthe aforementioned gradient elution treatment can be conducted with asecond solvent including the aforementioned ester solution and/or alkanesolution. In one example, the ester solvent of the second solvent can beethyl acetate, and the alkane solvent of the second solvent can ben-hexane. A concentration gradient (volume ratio) of n-hexane to ethylacetate of the second solvent for the gradient elution treatment can be100:0 to 0:100, but 80:20 to 40:60 are preferred.

In some specific examples, an n-hexane to the ethyl acetateconcentration gradient (volume ratio) of the second solvent can be 60:40to 50:50, for example, for the gradient elution treatment conducted onthe crude extract of the B. violaceum. In other specific examples, theconcentration gradient (volume ratio) of n-hexane to ethyl acetate ofthe second solvent can be 80:20 to 70:30, for example, for the gradientelution treatment conducted on the crude extract obtained from B.excavatum. In other specific example, the concentration gradient (volumeratio) of the n-hexane and the ethyl acetate of the second solvent canbe 70:30, for example, for the gradient elution treatment conducted onthe crude extract of the B. stechei.

It should be supplemented that, in other embodiments, a lipid removaltreatment can be optionally conducted after the crude extraction step(STEP 103 in FIG. 1) and/or together with the process of the columnchromatography step (STEP 107 in FIG. 1) in the method 100 of producingthe coral composite extract of the present invention. In theaforementioned example that the lipid removal treatment is conductedafter the crude extraction step (STEP 103 in FIG. 1), the crude extractcan be cold soaked in the ester solvent (for example, ethyl acetate) forprecipitating lipid with the ester solvent so as to remove most of thelipid in the crude extract. On the other hand, in the example that thelipid removal treatment is conducted together with the columnchromatography step (STEP 107 in FIG. 1), the crude extract can befilled in the chromatography column and be eluted with alkane solvent(such as n-hexane) and/or ester solvent (such as ethyl acetate) withlower polarities for the gradient elution treatment to remove most ofthe lipid in the crude extract, in which the volume ratio of the alkanesolvent (such as n-hexane) and ester solvent (such as ethyl acetate) canbe 100:0 to 95:5, for example.

In some embodiments, each aforementioned fractions obtained includingthe coral composite extract can be separated or combined together, andthe second solvent therein is further remove by the known dehydrationmethod (for example, vacuum concentration, lyophilization, spray drying,etc.) to obtain the coral composite extraction. Then, the obtained coralcomposite extract is found to have at least two briarane-typediterpenoid components, for example, EXC-B, EXC-C, EXC-X, BR, BR-X andBR-W, by analyzing with the known analytical method e.g., thin-layerchromatography (TLC), nuclear magnetic resonance spectroscopy (NMR),infrared spectroscopy (IR) and mass spectroscopy (MS), and so on.

It should be supplemented that, the obtained coral composite extractwill not contain the at least two briarane-type aforementionedditerpenoid compounds if the starting coral material is not originatedfrom the abovementioned specific species, or if the starting coralmaterial is not extracted with the abovementioned crude extraction stepand the column chromatography step.

In application, the abovementioned coral composite extraction can beapplied as an effective ingredient in compositions, i.e., a skinexternal use composition, a cosmetic composition and a medicinalcomposition, etc. In one example, the aforementioned composition can be,for example, applied to the test part by smearing or covering, etc., andthe formulation can be an emulsion, cream, hydrogel, gel, gel dressings,foam dressings, film dressings or any combination from above. Inapplication, the effective dose of the coral composite extract in theaforementioned composition can be 0.001 mg/mL to 20 mg/mL, for example.

It will be apparent to those skilled in the art that variousmodifications and variations can be made to the structure of the presentinvention without departing from the scope or spirit of the invention.In view of the foregoing, it is intended that the present inventioncover modifications and variations of this invention provided they fallwithin the scope of the followed claims.

Example 1: Preparing Coral Composite Extract

1. Coral Composite Extract Prepared from Briareum violaceum

In the present embodiment, Briareum violaceum (BV) was applied as thecoral stating material to obtain the coral composite extract. FIG. 2showed the partial flow chart of the method 200 of producing coralcomposite extract according to another embodiment of the presentinvention. Firstly, as shown in STEP 201, the lyophilized material ofthe BV coral was provided, in which the original wet weight of the BVcoral material was 231 g, and its lyophilized dry weight was 95 g. Next,as shown in STEP 203, the crude extraction step was performed on thelyophilized BV coral material by repeating the solvent extractiontreatment with ethyl acetate (1000 mL) as the first solvent for 20times, thereby obtaining the crude extract (3.3 g) in the ethyl acetatelayer. Then, as shown in STEP 207, the column chromatography step wasperformed on the crude extract (i.e., the ethyl acetate layer) byconducting the gradient elution treatment with a commercial availablechromatography column [in which the packing was silica gel (70-230 mesh,Merck)] and a second solvent as an eluent to obtain 7 fractions BV1 toBV7, in which the second solvent was a mixture of n-hexane and ethylacetate with a concentration gradient (volume ratio) of 100:0 (i.e.,pure n-hexane) to 95:5 (BV1, 0.11 g), 90:10 (BV2, 0.55 g), 80:20 (BV3,0.20 g), 70:30 (BV4, 0.23 g), 60:40 (BV5, 0.83 g), 50:50 (BV6, 0.22 g)and 40:60 to 0:100 (i.e., pure ethyl acetate) (BV7, 1.04 g).

By analyzing the aforementioned fractions with TLC, NMR, IR, MS, and soon, it was found that fraction BV1 included lipid, and the fraction BV6and the fraction BV7 included the coral composite extract, in which thefraction BV6 included 0.02 g of EXC-Z, while the fraction BV7 included0.6 g of EXC-B, as shown in STEP 209.

2. Coral Composite Extract Prepared from B. excavatum

In the embodiment, B. excavatum (BE) was applied as the coral startingmaterial to obtain the coral composite extract. FIG. 3 showed thepartial flow chart of the method 300 of producing the coral compositeextract according to the other embodiment of the present invention.Firstly, as shown in STEP 301, the lyophilized BE coral material wasprovided, in which the lyophilized dry weight of the BE coral materialwas 1900 g. Next, as shown in STEP 303, the crude extraction step wasperformed on the lyophilized BE coral with ethyl acetate (2000 mL) asthe first solvent for 20 times, in which the crude extraction stepincluded the solvent extraction treatment or the solvent extractiontreatment in combination with the ultrasonic treatment, therebyobtaining the crude extract (96.1 g) from the ethyl acetate layer.

Then, the crude extract (96.1 g) was cold soaked in ethyl acetate (800mL) for precipitating lipid so as to remove most of the lipid (5.6 g) inthe crude extract. After that, as shown in STEP 307, the columnchromatography step was performed on the crude extract (i.e., the ethylacetate layer, 90.5 g) with the same aforementioned commercial availablechromatography column for conducting the gradient elution treatment onthe crude extract with a second solvent as the eluent so as to obtain 8fractions BE1 to BE8, in which the second solvent was a mixture ofn-hexane and ethyl acetate with concentration gradient (volume ratio) of100:0 (i.e., pure n-hexane) (BE1), 95:5 (BE2), 90:10 (BE3), 80:20 (BE4),70:30 (BE5), 60:40 (BE6), 50:50 (BE7) and 40:60 to 0:100 (i.e., pureethyl acetate) (BE8).

By analyzing the aforementioned fractions with TLC, NMR, IR, MS, and soon, it was found that the fraction BE1 and the fraction BE2 includedlipid, and the fraction BE4 and the fraction BE5 included the coralcomposite extraction, in which the fraction BE4 included 16.2 g ofEXC-B, while the fraction BE5 included 3.4 g of EXC-C, as shown in STEP309.

Moreover, to evaluate the extraction efficiency of different extractiontreatments, the aforementioned solvent extraction treatment couldoptionally apply with a commercially available stirring device, forexample, Thermo hot-plate stirrer, (i.e., direct solvent extraction),which mixed first solvent at 150 rpm in the room temperature for 2 hourscontinuously with the coral starting material, which was then filteredand repeatedly extracted for five times.

For the solvent extraction treatment combining with the ultrasonictreatment, commercial available ultrasonic devices, for example, anultrasonic device A (130 Watt Vibra-Cell Ultrasonics Processor, Cat.#VCX130, Sonics, Hsinchu, Taiwan, power consumption as 130 W, frequencyas 20 kHz and dissipated power as 100%) and an ultrasonic device B (Cat.#DC900H, Delta, Taichung, Taiwan, power as consumption 900 W, frequencyas 40 kHz and dissipated as power 100%) could be used. When using theultrasonic device A, a treatment so-called ultrasonic homogenizerextraction proceeded every 10 sec with 1 second of break in a durationof 2 hours on the coral starting material, which was then filtered andrepeatedly extracted with the first solvent for five times. When usingthe ultrasonic device B, a treatment so-called ultrasonic cleanerextraction proceeded in a duration of 2 hours on the coral startingmaterial, which was then filtered and repeatedly extracted with thefirst solvent for five times. The table below showed the extractionrates of the aforementioned crude extraction step in different ways:

TABLE 1 Ultrasonic Ultrasonic Direct homogenizer cleaner solventextraction extraction extraction Dry weights of the 300.15 g 300.66 g300.39 g starting material Solid to liquid ratio 1:10 1:10 1:10(weight:volume) Ethyl acetate layer  6.48%  6.20%  6.18% extraction rateResidue recovery 90.96% 92.32% 90.11% rate

Please refer to TABLE 1 and FIG. 5, which showed the extraction rate(TABLE 1) of the crude extract obtained by the crude extraction step indifferent ways and the bar chart thereof (FIG. 5). In FIG. 5, thevertical axial represented the extraction rate (%) of the obtained crudeextract with the lyophilized dry weight considered as 100%.

Referring to Table 1 and FIG. 5, the solvent extraction treatment incombination with the ultrasonic treatment could indeed increase theextraction rate of the crude extract. Moreover, the extraction rate ofthe ultrasonic homogenizer extraction was higher than that of theultrasonic cleaner extraction and the direct solvent extraction.

3. Coral Composite Extract Prepared from B. stechei

In the embodiment, B. stechei (BS) was selected as the coral startingmaterial to obtain the coral composite extract. FIG. 4 was the partialflow chart of the method 400 of producing coral composite extractaccording to the other embodiment of the present invention. Firstly, asshown in STEP 401, the lyophilized BS coral material was provided, inwhich the original wet weight of the BS coral material is 6320 g, andits lyophilized dry weight was 2780 g. Next, as shown in STEP 403, thecrude extraction step was performed on the lyophilized BS coral materialwith ethyl acetate (3000 mL) as the first solvent for repeating thesolvent extraction treatment for 20 times, thereby obtaining the crudeextract (126.96 g) in the ethyl acetate layer. Then, the columnchromatography step was performed on the crude extract (i.e., ethylacetate layer) by performing the gradient elution treatment with theaforementioned commercial available chromatography column and the secondsolvent as the eluent on the crude extract to obtain 8 fractions BS1 toBS8, in which the second solvent was a mixture of n-hexane and ethylacetate with the concentration gradient (volume ratio) of 100:0 (i.e.,pure n-hexane) to 95:5 (BS1, 42.09 g), 90:10 to 80:20 (BS2, 3.81 g),70:30 to 60:40 (BS3, 4.63 g), 50:50 to 40:60 (BS4, 14.71 g), 30:70 to0:100 (i.e., pure ethyl acetate) (BE5, 25.89 g), 90:10 to 70:30 (BS6,2.33 g), 50:50 (BS7, 7.23 g) and 30:70 to 0:100 (i.e., pure ethylacetate) (BS8, 24.78 g).

By analyzing with TLC, NMR, IR, MS, and so on, it was found that thefraction BS1 and the fraction BS2 included lipid and the fraction BS5included the coral composite extract containing 3.79 g BR (as shown informula 1), 0.01 g BR-X (as shown in formula 2) and 0.02 g (shown informula 3).

It should be supplemented that, the coral composite extract in thisinvention can be applied as an effective ingredient of compositions, forexample, a skin external use composition, a cosmetic composition and amedicinal composition, etc.

In sum, although specific species of the corals, specific extractionmethods and specific evaluation methods are shown in the presentinvention as examples to explain the coral composite extract, thecomposition including the same and the method of producing the same, itwill be apparent to those skilled in the art that the present inventionis not limited to what have mentioned. Without departing from the scopeor spirit of the invention, it is intended that the present inventioncovers other coral species, methods for extraction and evaluation.

From the aforementioned embodiments, the advantages of the coralcomposite extract, the compositions including the same and the methodsof producing the same are that the coral composite extract contains atleast two briarane-type diterpenoid compounds and can be added to thecomposition as an effective ingredient.

Although the present invention has been described in considerable detailwith reference to certain embodiments thereof, other embodiments arepossible. Therefore, the spirit and scope of the appended claims shouldnot be limited to the description of the embodiments contained herein.

What is claimed is:
 1. A coral composite extract obtained from aBriareum excavatum coral starting material by (i) a crude extractionstep and (ii) a column chromatography step, wherein the crude extractionstep comprises a solvent extraction treatment combined with anultrasonic treatment, wherein the coral composite extract comprises atleast two briarane-type diterpenoid compounds that are excavatolide B(EXC-B) and EXC-C, and wherein the two briarane-type diterpenoidcompounds are the two that are present in the highest amounts in thecoral composite extract.
 2. A composition comprising the coral compositeextract of claim 1, wherein the coral composite extract is atherapeutically effective ingredient.
 3. The composition of claim 2,wherein the composition is a cosmetic composition or a medicinalcomposition.